Hypospadias is a congenital defect characterized by an ectopic opening of the urethra on the ventral side of the penis. It is one of the most common birth defects in males with an incidence of 1 in 125 live births. Currently, the only treatment for hypospadias is surgery. The high incidence of hypospadias is reflective of its multiple causes. Studies have found that environmental exposure of embryos to endocrine disrupting chemicals increases the rate of hypospadias. Additionally, there are syndromic forms of hypospadias that result from genetic mutations. However, in spite of recent progress in understanding external genitalia development it is still not clear exactly how genetic or environmental signals are influencing external genitalia development. The urethra is formed from a plate of endodermally derived epithelium that is surrounded by the mesoderm of the developing external genitalia, or genital tubercle (GT). The urethral plate epithelium cavitates to form the urethral tube, and failure to maintain the integrity of this tube leads to hypospadias. During development of the GT, an ectodermal opening at the base of the GT exposes the underlying urethral tube and occurs in both male and female embryos, but this proximal urethral opening (puo) is closed at later stages in males while it is maintained in females to become the vaginal opening. Interestingly, many cases of hypospadias occur in the proximal penis leading to the hypothesis that defective morphogenesis of the puo causes proximal hypospadias. To address this hypothesis, this proposal will utilize three complimentary approaches. Aim 1 of this proposal will examine the cellular events that control normal formation and closure of the puo during development of the male mouse GT. Aim 2 will address molecular control of puo morphogenesis through examination of changes in gene expression and cellular events in Bmp2 and Fgfr2 mutants. Both Bmp2 and Fgfr2 are expressed in the puo region, and Fgfr2 mutants show defective puo morphogenesis. Finally, Aim 3 will examine the cellular and molecular mechanisms that lead to the naturally occurring open urethra in the red-eared slider turtle Trachemys scripta. This aim will allow for the identification of novel mechanisms important for maintaining urethral closure. Findings from this proposal will not only add to the knowledge of how external genitalia normally develop, but will also identify mechanisms that lead to the generation of hypospadias. This could have broader implications in the development of methods to prevent hypospadias. Additionally, work in T. scripta will aid in determining factors that are involved in the generatio of hypospadias and may lead to a novel model system for studying hypospadias in ovo. PUBLIC HEALTH RELEVANCE: Hypospadias, a defect in urethral tube closure, is one of the most common birth defects occurring in 1 in 125 male births; however, the cause of hypospadias is not clear. This proposal will examine morphogenesis of the proximal urethral opening, a structure that forms in both males and females, but is normally closed at later stages in males. As defective closure of the proximal urethral opening would lead to hypospadias in males, this research will provide insight into the molecular and cellular mechanisms that lead to hypospadias in the proximal urethra.